Despite recent progress in treatment and prevention of coronary heart disease, sudden cardiac death (SCD) remains a major public health problem, with an annual incidence of 180,000-250,000 in the U.S. The vast majority of SCD events occur in the general population, with up to 50% of individuals experiencing SCD as a first sign of disease. The critical importance of genetic contribution for effective prediction and prevention of SCD was emphasized in a recent consensus document from the National Heart Lung and Blood Institute. We have recently identified a region of the genome, designated SCD1, containing three genes, BAZ2B, WDSUB1 and TANC1, that is strongly associated with risk for SCD (P=2.2x10-11). The risk allele, while relatively rare (~3% of individuals of European ancestry are carriers), has a large effect, increasing the risk for SCD by 2.03-fold (95% CI 1.65-2.49). We hypothesize that one or more of the genes in the SCD1 region contribute to cardiovascular development and function and that coding or noncoding variation therein plays an important role in SCD risk. First, we will evaluate the biological relevance of each of the 3 genes using both mouse and zebrafish model organisms. We will determine the spatial and temporal distribution of the transcripts across a range of developmental and post-natal stages in mice through both whole mount RNA in situ analyses and sectioning of embryonic and postnatal heart. We will use zebrafish to test the hypothesis that disruption of the expression of one or more gene candidates during development of vertebrate organism will compromise the genesis and function of cardiovascular components. We anticipate that these experiments will definitively implicate one or more of the genes in cardiovascular biology, and further, provide preliminary evidence for a biological mechanism by which local genetic variation alters the risk for SCD in humans. Second, we will characterize the genetic variation in this region in 3,900 autopsy-identified SCD cases and >6,000 controls to: 1) identify the functional variant(s) underlying the identified association signal; 2) identify additional variants associated with SCD; 3) dissect the nature of the genetic association using detailed autopsy data to define the cause of SCD and stratify samples. We anticipate that both non-coding and coding variants will be associated with SCD. For coding variation, we will compare the capacities of human RNAs containing identified coding variation with their non-variant counterparts to rescue MO-induced effects. We will similarly assay the effects of over-expression. For noncoding variation, we will first assay the regulatory control of selected associated noncoding sequences by transgenesis in zebrafish. Additionally we will determine whether identified variants alter endogenous gene expression levels (eQTL) of RNA obtained from the autopsy hearts. This application bridges a critical gap between genetic association and functional studies, leveraging a unique autopsy-determined SCD repository, high-throughput next generation sequencing, and model organisms to functionally and genetically dissect a highly significant finding from a GWAS. PUBLIC HEALTH RELEVANCE: The overall goal of this proposal is to functionally and genetically characterize a region of the genome that has been associated with risk of sudden cardiac death (SCD). SCD is one of the leading causes of mortality in the United States, and given the lack of predictive markers for the vast majority of SCD events, which occur in the setting of subclinical heart disease and have >90% mortality, the proposed studies are critical for both risk stratification and to identify specific therapeutic targets.